The oxidized and reduced forms of the four-iron ferredoxin from the hyperthermophilic archaeon, Thermococcus litoralis (Tl), have been investigated by H-1 NMR spectroscopy. All three protons for each of the four Cys are located by 2D and 1D NMR experiments, although differentiation of alpha- and beta-protons required the interpretation of steady-state NOEs and relaxation properties of the signals. The complete correlation between the Cys signals in the reduced and oxidized ferredoxin was carried out by 2D EXSY and steady-state saturation-transfer NMR, and an electron self-exchange rate of similar to 5 x 10(4) M(-1) s(-1) at 30 degrees C, pH 7.6, is estimated. The beta-protons for two ligated Cys exhibit Curie-like temperature dependence, and the other two display anti-Curie temperature dependence, indicative of the iron atoms arising from the mixed-valence pair with intermediate S = 9/2, and the diferrous pair with intermediate S = 4, respectively. Standard 2D identification of the protons for nonligated residues which have significant dipolar contacts with the ligated Cys provide the sequence-specific assignment for Cys 18 and Cys 16. The two remaining ligated Cys are shown to be sequence-specifically assignable upon qualitative consideration of the relative relaxation properties of the ligated Cys alpha-protons, as predicted by the pattern of Cys orientations relative to the cluster in numerous crystallographically characterized bacterial ferredoxins. The combined assignments identify Cys 10 and Cys 16 as ligated to the valence-delocalized iron atoms in the reduced ferredoxin. Comparison of the NMR spectra of the reduced four-iron and oxidized three-iron Tl ferredoxins indicates a similar pairing of the iron atoms in the spin coupling hierarchy. The patterns of both relaxation properties and contact shifts of alpha-protons for ligated Cys are discussed in terms of the geometry of the ligand.